Genetic engineering now provides the tools to begin to develop herbicide resistance crops, thereby offering valuable options in effective crop management. The ability to introduce herbicide resistance into plants extends the use of desirable, non-selective herbicides to sensitive crop species, and offers numerous economic and environmental advantages (Schulz et al., 1990). An excellent example of this is the commercial introduction of glyphosate resistance in a variety of crops.
The pyridine family of herbicides can control narrow-leaf and small seeded broad-leaf weeds under pre-emergence application (See for example, U.S. Pat. Nos. 4,826,532; 4,747,871; 4,692,184; 4,885,026; and 5,019,153). Cell biology studies using the pyridine dithiopyr suggest that disruption of cell division is the mode of action of the pyridine herbicides (Armbruster et al., 1991), most likely by disrupting microtubule organization (Armbruster et al., 1988). Seeds germinated in the presence of pyridine herbicides show characteristic inhibition of root elongation and swelling of meristematic zones. Thiazopyr is predominantly metabolized in both animals (Feng et al., 1994a) and plants (Feng et al., 1995a) via oxidations at the sulfur or carbon atoms in the thiazoline ring. The resulting initial metabolites have a transient existence and are further degraded to polar and/or acidic metabolites (McClanahan et al., 1995). A key reaction is de-esterification of the methylester functional group to form the monoacid metabolite which is virtually devoid of any herbicidal activity (Feng et al., 1995b). It has been demonstrated in animals that de-esterification can occur either by oxidation (Feng et al, 1994b) or hydrolysis (Feng et al., 1995b), both reactions being catalyzed by liver enzymes (Feng et al. (1994b)). Feng et al. (1995b) were particularly interested in esterases due to their lack of requirement for cofactors during catalysis and their ubiquitous presence in nature hydrolyzing both endogenous substrates and xenobiotics (Leinweber, 1987). Thiazopyr metabolism in plants differs from animals in that the monoacid appears to be produced exclusively via the oxidation pathway (Feng et al., 1995). Using inhibitors of monooxygenase enzymes, Feng et al. (1995a) demonstrated in planta suppression of thiazopyr metabolism which translated to enhanced bioefficacy.
Thiazopyr shows little plant selectivity which limits its use in many important agronomic crops. The present application describes the purification of a novel thiazopyr esterase, and the subsequent cloning and expression of a gene encoding the esterase in plants. Tobacco and tomato plants transformed to express the pyridine-esterase confirm that esterase-mediated deactivation of pyridines is a viable approach for engineering herbicide resistance in plants.